開發肺部遞送之吸入性光敏劑微奈米乾粉

• Main Author: 楊育才
• Other Authors: 牙醫學系碩博士班
• Language: Chinese; English
• Published: 2010
• Subjects: Angeli; Birchall; Dougherty; Ferro; Konan; Leroux; Lier; McCarthy; Perez; Roa; St. Louis; Williamson; laptev
• Online Access: http://libir.tmu.edu.tw/handle/987654321/36289

Content.1 Table Captions.4 Figure Captions.5 摘 要.8 Abstract.10 Chapter 1 Introduction.12 Chapter 2 Literature Review.17 2.1 Photodynamic therapy.17 2.2 Photosensitizer.18 2.3 Nanocarrier encapsulating photosensitizers.19 2.4 Nanocarriers.20 2.4.1 Liposomes.21 2.4.2 Polymeric micelles.22 2.4.3 Nanoparticles.23 2.5 Pulmonary inhalation systems.23 2.5.1 Effect of particle sizes on aerosol deposition.26 Chapter 3 Materials and Methods.27 3.1 Preparation and characterization of Hp encapsulated in nanocarriers.27 3.1.1 Preparation of Hp encapsulated in liposomes.28 3.1.2 Preparation of Hp encapsulated in micelles.29 3.1.3 Preparation of Hp encapsulated in nanoparticles.29 3.1.4 Characterization of nanocarrier Hp.30 3.1.5 Cell culture testing.34 3.2 Preparation and characterization of microparticles containing micelles encapsulating Hp.37 3.2.1 Preparation of micellar Hp within lactose microparticle.37 3.2.2 Characterization of micellar Hp within lactose microparticle.38 3.2.3 Cell culture testing.40 Chapter 4 Results.44 4.1 Characterization of nanocarrier Hp.44 4.1.1 Size of nanocarrier Hp.44 4.1.2 Photophysical characterization of Hp in aqueous, ethanol, and nanocarrier.44 4.1.3 Photophysical characterization of Hp in liposomes.47 4.1.4 Photophysical characterization of Hp in micelles.48 4.1.5 Photophysical characterization of Hp in nanoparticles.49 4.1.6 Singlet oxygen generation of Hp in nanocarriers.49 4.1.7 In vitro Hp release.50 4.2 Cellular uptake and photocytotoxicity of free Hp and nanocarrier Hp.51 4.3 Characterization of Hp encapsulated in micelles within the microparticle powder.52 4.3.1 Drug loading and entrapment efficiency of micellar Hp.52 4.3.2 Size and spectrum of micellar Hp before and after spray-drying.53 4.3.3 Singlet oxygen generation of free Hp, micellar Hp and lactose-micellar Hp.54 4.3.4 Optical and fluorescence microscope images of lactose-micellar Hp.55 4.3.5 Size and morphology of lactose-micellar Hp.55 4.4 Intracellular distribution of Hp fluorescence in A549 tumor cells.56 4.5 Photocytotoxicity of free Hp, micellar Hp, lactose-micellar Hp.56 Chapter 5 Discussions.58 5.1 Spectrum study of Hp encapsulated in nanocarriers.58 5.2 Microparticles containing micellar Hp.63 Chapter 6 Conclusions.68 6.1 Hp encapsulated in nanocarrier systems.68 6.2 Microparticles containing micellar Hp.68 References.70 Publications.127 1.Lambert CR, Reddi E, Spikes JD, Rodgers MAJ, Jori G. 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J Photochem Photobiol B 2002;66(2):89-106. 本研究目的為開發新穎之吸入性光敏劑微奈米乾粉，一種結合奈 米與微米複合劑型並包覆光敏劑血紫質(hematoporphyrin, Hp)之肺部遞送系統。本研究以吸收及螢光光譜分析方式監測血紫質於奈米包埋系統(微脂體、微胞、奈米粒)中，以最佳化的單體分散形式存在，避免因分子聚集而造成光動力效應的功效損失。研究發現微脂體、微胞、以及奈米粒這三種奈米載體均可有效降低水不溶性光敏劑Hp 的分子聚集，也均有提高光動力之效應。本研究進一步以人類肺腺癌細胞株A549 作體外模式細胞試驗，評估三種奈米包埋系統包覆光敏劑血紫質細胞之攝入能力及所引發的光動力效應。結果顯示，微胞化之光敏劑血紫質光動力效應明顯優於微脂體及奈米粒系統。因此，選擇光動力效應最高微胞化光敏劑血紫質，以噴霧乾燥之方式結合乳糖乾粉微粒包埋光敏劑血紫質微胞。藉由吸收光譜分析、單態氧產生及粒徑分析結果，噴霧乾燥後的微胞仍維持噴霧前之物化性質，且藉由電子顯微鏡結果分析微粒乾粉粒徑為2.3 ± 0.7 μm，此粒徑範圍可有效沉積肺泡。本實驗並以A549 作體外模式細胞試驗，結果也顯示出噴霧前後微胞包覆光敏劑血紫質細胞攝入能力及所引發的光動力效應皆無明顯差異，但皆明顯優於未包覆之組別。因此利用噴霧乾燥之方式結合乾粉微米及微胞為一極具潛力之肺部給藥系統。 The purpose of this study was to examine the properties of a new pulmonary delivery platform of microparticles containing nanocarriers in which a therapeutic photosensitizing drug, hematoporphyrin (Hp), was entrapped. Different nanocarriers including liposome, micelle and nanoparticle were used. Absorption and emission spectral analysis were used to monitor the monomeric/aggregated state of Hp in different carriers and solvent systems. The high levels of monomer Hp in the DMPC liposome, L122 micelle and PLGA nanoparticle was observed in all formulation. One of these, Hp encapsulated in L122 micelle (micellar Hp), was subsequently incorporated into lactose microparticles by spray-drying due to highest cellular uptake and photocytoxicity in human lung epithelial carcinoma A549 cells. Spectral and morphological analyses were performed on both micellar Hp, and lactose microparticles containing micellar Hp (lactose-micellar Hp) before and after dissolution of the microparticles in water. Photodynamic activity of the various Hp samples were evaluated in A549 cells using a light emitting diode (LED) device at a wavelength of 630 ± 5 nm. It was found that there were no significant differences between micellar Hp and lactose-micellar Hp on the generation of singlet oxygen. The mean particle size of the microparticles was 2.3 ± 0.7 μm which is within the sizerange for potential lung delivery. The cellular uptake of micellar Hp and lactose-micellar Hp measured on A549 cells was at least two-fold higher than those obtained with the Hp at equivalent concentrations. Micellar Hp exhibited higher cytotoxicity than Hp due to reduced formation of Hp aggregates and increased the cellar uptake. The spectral properties as well as the photodynamic activity of the micellar Hp was retained when formulated into microparticles by spray-drying. Microparticles containing micelles has the potential to deliver the micelle-encapsulated hydrophobic drugs for targeted therapy of pulmonary diseases.